Notch Signaling Pathway Was Involved In Regulating Programmed Cell Death 1 Expression During Sepsis-induced Immunosuppression

?Objective?Immunosuppression is one of main factors which influence the mortality of sepsis.PD-1 and its ligands PD-L1 and PD-L2 is a kind of co-inhibitory molecules and the activation of PD-1 signaling pathway can regulate the immune system negatively.Animal experiments had demonstrated that blocking the PD-1 signaling pathway via administration of PD-1 or PD-L1 antibody can improve the survival of septic mice by inhibiting the lymphocyte apoptosis and reversing monocyte dysfunction.In our research,we first clarify the relationship between PD-1,PD-L1 expression and the severity and prognosis of septic shock patients.Then,we explore the mechanism underlying the regulation of PD-1 expression to provide new insights into the immune therapy of sepsis.?Methods?(1)clinical studyPatients satisfying the definition of septic shock were included and peripheral blood was collected within 24h after diagnosis of septic shock.Healthy volunteers matched with sex and age were enrolled as control group.To measure the expression of PD-1 and PD-L1 on the CD4~+T lymphocyte and CD14~+monocyte by flow cytometry and compare the level of them between the septic shock patients and healthy volunteers or survivals and non-survivals.To isolate the peripheral blood mononuclear cells(PBMCs)and measure the expression of Notch1,Jagged1 and Hes1 by real-time PCR and compare the level of them between the septic shock patients and healthy volunteers or survivals and non-survivals.(2)experiments in vitroEndotoxin tolerance cell model was set up and divided into four groups:control group,LPS non-tolerant group,LPS tolerant group and LPS tolerant+DAPT group.To measure the mRNA levels of TNF-?,IL-10,PD-1,Jagged1 and Hes1 by real-time PCR and the expression of PD-1 on THP1 cells by flow cytometry and compare the levels of TNF-?,IL-10,PD-1,Jagged1 and Hes1 among groups.?Results?(1)clinical study1.Compared with healthy volunteers,the level of PD-1 and PD-L1 expression on CD4~+T lymphocyte and CD14~+monocyte in septic shock patients were significantly higher(CD4~+T lymphocyte:21.59%vs 6.37%,p<0.001;13.67%vs 4.97%,p<0.01;CD14~+monocyte:14.91%vs 6.89%,p<0.01;51.82%vs 17.12%,p<0.01).The level of PD-1 and PD-L1 on CD14~+monocyte were correlated positively with the APACHEII score(r=0.52,p<0.0001;r=0.55,p<0.0001).Compared with survivors,the levels of PD-1 and PD-L1expression in non-survivors were significantly higher(19.13%vs 13.79%,p<0.05;60.21%vs 45.06%,p<0.01).2.Compared with healthy volunteers,the mRNA levels of Jagged1 and Hes1 in septic shock patients were significantly higher.Compared with survivors,the mRNA levels of Jagged1 in non-survivors were significantly higher.(2)experiments in vitro1.Compared with LPS non-tolerant group,the levels of IL-10,Jagged1,Hes1 and PD-1 in LPS tolerant group were significantly higher,while the level of TNF-?was significantly lower.2.The levels of PD-1 and IL-10 were significantly lower in the group treated with DAPT when compared with non-treatment group.?Conclusions?1.The level of PD-1 and PD-L1 was elevated in septic shock patients and associated with the severity and prognosis of sepsis.2.Notch signaling pathway was activated in patients at early stage of septic shock.3.Notch signaling pathway was involved in regulating PD-1 expression during sepsis-induced immunosuppression.